G protein coupled receptors (GPCRs) were first characterized as signal transducers that elicit downstream effects through modulation of guanine (G) nucleotide-binding proteins. The pharmacotherapeutic exploitation of this signaling paradigm has created a drug-based field covering nearly 50% of the current pharmacopeia. Since the groundbreaking discoveries of the late 1990s to the present day, it is now clear however that GPCRs can also generate productive signaling cascades through the modulation of β-arrestin functionality. β-Arrestins were first thought to only regulate receptor desensitization and internalization – exemplified by the action of visual arrestin with respect to rhodopsin desensitization. Nearly 20 years ago, it was found that rather than controlling GPCR signal termination, productive β-arrestin dependent GPCR signaling paradigms were highly dependent on multi-protein complex formation and generated long-lasting cellular effects, in contrast to G protein signaling which is transient and functions through soluble second messenger systems. β-Arrestin signaling was then first shown to activate mitogen activated protein kinase signaling in a G protein-independent manner and eventually initiate protein transcription – thus controlling expression patterns of downstream proteins. While the possibility of developing β-arrestin biased or functionally selective ligands is now being investigated, no additional research has been performed on its possible contextual specificity in treating age-related disorders. The ability of β-arrestin-dependent signaling to control complex and multidimensional protein expression patterns makes this therapeutic strategy feasible, as treating complex age-related disorders will likely require therapeutics that can exert network-level efficacy profiles. It is our understanding that therapeutically targeting G protein-independent effectors such as β-arrestin will aid in the development of precision medicines with tailored efficacy profiles for disease/age-specific contextualities.

SummaryDesensitization, signaling and trafficking of G protein-coupled receptors (GPCRs) are critically regulated by multifunctional adaptor proteins, β-arrestins (βarrs). The two isoforms of βarrs (βarr1 and 2) share a high degree of sequence and structural similarity, still however, they often mediate distinct functional outcomes in the context of GPCR signaling and regulation. A mechanistic basis for such a functional divergence of βarr isoforms is still lacking. Using a set of complementary approaches including antibody fragment based conformational sensors, we discover structural differences between βarr1 and 2 upon their interaction with activated and phosphorylated receptors. Interestingly, domain swapped chimeras of βarrs display robust complementation in functional assays thereby, linking the structural differences between the receptor-bound βarr1 and 2 with their divergent functional outcomes. Our findings reveal important insights into the ability of βarr isoforms to drive distinct functional outcomes, and underscore the importance of integrating this aspect in the current framework of biased agonism.

Highlights d b-arrestin 1 and 2 differentially regulate signaling and trafficking of GPCRs d There are conformational differences between receptorbound b-arrestin 1 and 2 d Domain-swapped chimera of b-arrestin 1 and 2 exhibit functional complementation d Functional differences of b-arrestin isoforms have implications for biased agonism